Devices, systems, and methods for targeted plating of materials in high-throughput culture plates

ABSTRACT

Devices, systems, and methods for facilitating placement of cells and materials in culture plates configured for high-throughput applications are provided. A culture system is provided with a culture plate having a lid for guiding placement of cells and materials in each individual culture well of a culture plate. The lid may provide for coupling to an electrophysiology culture plate comprising a biosensor plate and a biologic culture plate, where the biosensor plate underlies and is coupled to the culture well plate such that each biosensor is operatively coupled to one culture well of the plurality of culture wells. A containment device that physically influences the positioning of fluid received in the culture plate is also provided herein.

This application is a divisional of U.S. patent application Ser. No.14/533,373, filed on Nov. 5, 2014, which claims the benefit of U.S.Provisional Patent Application No. 61/899,970, filed on Nov. 5, 2013,the disclosures of which are expressly incorporated herein by referencein their entireties.

BACKGROUND OF THE INVENTION

Implementations described herein relate generally to devices, systemsand methods enabling targeted plating of materials in high-throughputculture plates, and, more particularly, to culture well plates and lidsconfigured to facilitate targeted placement of materials into anindividual culture well.

In vitro electrophysiology culture systems having biosensors, such asmicroelectrode arrays (MEAs), can provide important insights intonetworks of electrically active cells. MEA-based electrophysiologyculture systems can be configured to concurrently monitor single-celland network-level activity over extended periods of time and withoutaffecting the cell culture under investigation. Since their instrumentalrole in the landmark discovery of spontaneous waves in a developingretina, the variety and scope of MEA-based electrophysiologyapplications has dramatically expanded. Recently, for example, MEA-basedelectrophysiology culture systems have been used to investigate thesuppression of epileptic activity and in the study of novel plasticitymechanisms in cultured neural networks. Advances in cell culturepreparations have similarly led to applications for MEA-basedelectrophysiology culture systems in the fields of drug screening,safety pharmacology, and biosensing.

Present day MEA-based electrophysiology culture systems are typicallydesigned around small-footprint, single-well devices. However, thecomplete analysis of complex cellular systems and processes can requirerepeated experiments. The number of experiments can increase quicklywhen considering multiple variables, such as, for example and withoutlimitation, stimulus size, compound type, dosage strength and the like.Thus, the small-scale format of traditional MEA systems presents a“large N” problem (i.e., problems due to excessive experimental andstatistical sampling sizes), whereby the serial nature of these devicescan render even basic investigations time and cost prohibitive. As oneillustrative example, a researcher examining the effect of pythrethroidson two-hour spontaneous activity recordings can require 8 doses ofpermethrin, with an N of 6 for each dose. With traditional MEA-basedelectrophysiology culture systems, this very simple experiment canrequire over $5,000 in MEA-based electrophysiology culture plates (or“MEA culture plates”) and 50 to 60 man-hours. The time investment canfurther increase with the logistics of culturing, maintaining, andtesting dozens of individual specimen.

The applicant has developed high-throughput MEA culture plates in anANSI/SLAS compliant format to achieve industry compliance with otherhigh-throughput instrumentation such as robotic handlers and platereaders. Such MEA culture plates are described in U.S. ProvisionalPatent No. 61/899,970, filed on Nov. 5, 2013, entitled “Devices, Systemsand Methods for Targeted Plating Of Materials In High-Throughput CulturePlates,” which is hereby incorporated by reference in its entirety. Suchhigh-throughput culture plates can have well counts of, for example andwithout limitation, 12, 24, 48, 96, 192, 384 or 768. Further, each wellplate can have an area of interest, e.g. an electroactive area that canbe, for example and without limitation, about 1.25 mm to 2 mm indiameter.

SUMMARY

Plating cells and other materials (e.g., biomolecular coatings) canbecome a challenge when well counts are higher than about 12.Traditionally, materials are plated in such plates usingpipetting—either manually or automatically. Manual plating can becumbersome, time consuming, and inefficient. Automated plating withrobotic handlers can be more efficient but also more costly than manualplating. Thus, even if utilizing automated plating, time and money canbe saved by more accurately guiding the plating of cells and othermaterials. Accordingly, a need exists for improved devices, systems, andmethods that enable targeted plating of cells and other materials inhigh-throughput culture systems.

Implementations described herein are directed toward devices, systems,and methods for facilitating placement of cells and materials in cultureplates configured for high-throughput applications. Some implementationsare directed to a culture system comprising a culture plate having a lidconfigured to guide placement of cells and materials in each individualculture well of a culture plate. Some implementations are directedtoward culture wells with containment features or devices configured toconcentrate the volume of the cells and other materials to a biosensorarea.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a schematic of an example embodiment of an electrophysiologyculture plate and the associated electronics.

FIG. 2A is a schematic of an example embodiment of an electrophysiologyculture plate.

FIG. 2B is an exploded view of the electrophysiology culture plate fromFIG. 2A.

FIG. 3 is a step-by-step diagram of an example process of plating a cellsuspension in an electrophysiology culture plate.

FIG. 4 is a step-by-step diagram of an example process of plating of acell suspension in an electrophysiology culture plate guided by atargeted plating lid.

FIG. 5A shows a cross section of an example biologic culture plate lidwith various targeting devices for guiding the plating of a cellsuspension.

FIG. 5B shows a cross section of an example targeting device and apipette.

FIG. 6A is a top-down view of an example targeting device for guidingthe plating of a cell suspension.

FIG. 6B is a side view of an example targeting device.

FIG. 7A is a top-down view of an example targeting device for guidingthe plating of a cell suspension.

FIG. 7B is a side view of an example targeting device.

FIG. 8A is a top-down view of an example targeting device for guidingthe plating of a cell suspension.

FIG. 8B is a side view of an example targeting device.

FIG. 9A shows a cross section of an example biologic culture plate lidattached to the biologic culture plate with a plurality of targetingdevices.

FIG. 9B shows a cross section of an example lid fitting onto anelectrophysiology culture plate.

FIG. 10A shows a cross section of an example lid having a double baffle.

FIG. 10B is a perspective view of an example lid having a double baffleand a friction fit feature.

FIG. 10C is a perspective view of an example biologic culture plate lidhaving a friction fit feature.

FIG. 11 shows a top-down view of an example culture well with biosensorsand a mistargeted droplet.

FIG. 12A shows a top-down view of an example culture well withbiosensors and containment devices.

FIG. 12B shows a cross-sectional view along line A-A′ of FIG. 12A.

DETAILED DESCRIPTION

The present invention can be understood more readily by reference to thefollowing detailed description, examples, drawing, and claims, and theirprevious and following description. However, before the present devices,systems, and/or methods are disclosed and described, it is to beunderstood that this invention is not limited to the specific devices,systems, and/or methods disclosed unless otherwise specified, as suchcan, of course, vary. The terminology used herein is for the purpose ofdescribing particular aspects only and is not intended to be limiting.

The following description of the invention is provided as an enablingteaching of the invention in its best, currently known aspect. To thisend, those skilled in the relevant art will recognize and appreciatethat many changes can be made to the various aspects of the inventiondescribed herein, while still obtaining the beneficial results describedherein. It will also be apparent that some of the desired benefitsdescribed herein can be obtained by selecting some of the featuresdescribed herein without utilizing other features. Accordingly, thosewho work in the art will recognize that many modifications andadaptations to the present invention are possible and can even bedesirable in certain circumstances and are a part described herein.Thus, the following description is provided as illustrative of theprinciples described herein and not in limitation thereof.

Reference will be made to the drawings to describe various aspects ofone or more implementations of the invention. It is to be understoodthat the drawings are diagrammatic and schematic representations of oneor more implementations, and are not limiting of the present disclosure.Moreover, while various drawings are provided at a scale that isconsidered functional for one or more implementations, the drawings arenot necessarily drawn to scale for all contemplated implementations. Thedrawings thus represent an exemplary scale, but no inference should bedrawn from the drawings as to any required scale.

In the following description, numerous specific details are set forth inorder to provide a thorough understanding described herein. It will beobvious, however, to one skilled in the art that the present disclosuremay be practiced without these specific details. In other instances,well-known aspects of electrophysiology culture systems, machiningtechniques, injection molding methodologies, and microelectromechanicalsystems (MEMS) have not been described in particular detail in order toavoid unnecessarily obscuring aspects of the disclosed implementations.

Disclosed are components that can be used to perform the disclosedmethods and systems. These and other components are disclosed herein,and it is understood that when combinations, subsets, interactions,groups, etc. of these components are disclosed that while specificreference of each various individual and collective combinations andpermutation of these may not be explicitly disclosed, each isspecifically contemplated and described herein, for all methods andsystems. This applies to all aspects of this application including, butnot limited to, steps in disclosed methods. Thus, if there are a varietyof additional steps that can be predefined it is understood that each ofthese additional steps can be predefined with any specific aspect orcombination of aspects of the disclosed methods.

Implementations described herein are directed toward devices, systemsand methods for facilitating placement of cells and materials in cultureplates configured for high-throughput applications. More particularly,the present disclosure is directed to a culture system comprising aculture plate having a lid configured to guide placement of cells andmaterials in each individual culture well of a culture plate. Forexample, one or more implementations described herein provide for lidfor coupling to an electrophysiology culture plate comprising abiosensor (such as a MEA) array plate having a plurality of biosensorsand a biologic culture plate having a plurality of culture wells whereinthe biosensor plate underlies and is coupled to the culture well platesuch that each biosensor is operatively coupled to one culture well ofthe plurality of culture wells. As described herein, a biosensor may bea device used to measure electrical activity and to electricallystimulate a cell culture. For example, a biosensor may be an electrodeor a microelectrode. A biosensor may also measure the pH, ionicconcentration, impedance, strain, metabolite concentration, temperature,growth rate, proliferation rate, or any other measurable aspect of acell culture.

Reference will now be made to the drawings to describe various aspectsof one or more implementations of the invention. It is to be understoodthat the drawings are diagrammatic and schematic representations of oneor more implementations, and are not limiting of the present disclosure.Moreover, while various drawings are provided at a scale that isconsidered functional for one or more implementations, the drawings arenot necessarily drawn to scale for all contemplated implementations. Thedrawings thus represent an example scale, but no inference should bedrawn from the drawings as to any required scale.

High-throughput screening (HTS) tools make use of multi-well biologicculture plates that follow exacting guidelines established by theSociety for Lab Automation and Screening (SLAS) and the AmericanNational Standards Institute (ANSI). These standards are adhered to byall HTS supporting equipment such as, for example and withoutlimitation, plate readers, robotic handlers, liquid handling devices andthe like. Compliance with these standards can enable a high-throughputbiosensor platform to achieve full potential as it leverages existinghigh-throughput infrastructure including the automation of mediaexchanges and compound delivery.

In the following description, numerous specific details are set forth inorder to provide a thorough understanding described herein. It will beobvious, however, to one skilled in the art that the present disclosuremay be practiced without these specific details. In other instances,well-known aspects of electrophysiology culture systems machiningtechniques, injection molding methodologies and microelectromechanicalsystems (MEMS) have not been described in particular detail in order toavoid unnecessarily obscuring aspects of the disclosed implementations.Although a high-throughput electrophysiology culture system is describedbelow for clarity, the culture well plate lids described herein areapplicable to all other high-throughput culture plate systems.

Turning now to FIG. 1 in one embodiment, an electrophysiology cultureplate 102 can comprise a monolithic biosensor plate such as amicroelectrode array (MEA) plate 104 integrated with a biologic cultureplate 106, and electronics 108 together with software configured tostimulate a cell culture via the electrophysiology culture plate toevoke a response and to record data. The electrophysiology culture platecan comprise a plurality of culture wells 110 configured to cultureelectroactive cells. A grid of tightly spaced microelectrodes 112configured to extracellularly interface with the cultured cells may beoperatively associated with each culture well 110. Each microelectrodeor biosensor can be configured to record electrical activity from nearbyneurons and electrically stimulate those cells. This technique canprovide an extracellular, label-free method for examining bothindividual neuronal behavior and overall network activity, optionally,simultaneously. Mechanical features can also be provided that operate tocouple the biosensor culture well plate 102 to the electronics 108. In afurther aspect, these mechanical features can be configured so as not tointerfere with topside access to the electrophysiology culture plate. Inanother aspect, the electronics unit can be used amplify and filter thelow amplitude extracellular signals captured by both the microelectrodesand reference electrodes and, in other aspects, provide user directedstimulation to the cells. Reference electrodes can maintain apredetermined range or level of a given measurement, allowing forfiltering of background noise from the microelectrode measurement. Insome embodiments, a reference sensor may be a reference electrode thatmaintains a predetermined voltage range. For example, a reference sensormay be a ground electrode. In further aspects, the electronics unit canconvert the analog electrode signals received from the cells into datathat can be used and manipulated by the computer software, whileminimizing the amount of noise injected into the very low amplitudesignals that are measured (e.g., extracellular recordings).

The electrophysiology culture plate 102 shown in FIG. 2A can provide forhigh-throughput MEA culture well plates. FIG. 2B shows an exploded viewof the electrophysiology culture plate, including a monolithic MEA plate104 that can be fully vertically integrated with a culture well plate106. In a further aspect, the monolithic MEA plate 104 and culture wellplate 106 can be joined by an intermediate adhesive 114. The length andwidths of the MEA plate 104 and adhesive material 114 may be sized tomatch the dimensions of the biologic culture plate 106. The overall sizeof the culture plate 106 may vary depending on the number and sizes ofculture wells included on the plate. Any suitable size wells may beused, and any suitable culture plate size may be used as well. In someimplementations, culture plate dimensions adhere to ANSI/SLAS microplatestandards to achieve industry compliance with other high-throughputinstrumentation such as robotic handlers and plate readers. Accordingly,it is a further aspect of this invention to provide culture well platesdesigned to provide at least 1 culture well. In some embodiments, theculture well plate may include at least 2, 4, 6, 12, 24, 48, 96, 384,768, or more culture wells. In another aspect, a monolithic biosensorplate can comprise, for example and without limitation, polymers, glass,glass-reinforced epoxy resin, silicon, and the like.

In one aspect of the present disclosure, a multi-well biologic cultureplate configured to be attached to the biosensor plate can be provided.In light of the present disclosure, one skilled in the art willappreciate that multi-well biologic culture plates may includemicro-titer plates. In a further aspect, the biologic culture plate cancomprise a lid. The biologic culture plate and lid can be formed, forexample and without limitation, by conventional injection moldingtechniques. In a further aspect, the biologic culture plate and lid cancomprise materials such as, for example and without limitation,polystyrene, polycarbonate, cyclic olefin polymer, cyclic olefinco-polymer, and the like.

In other aspects, the present disclosure provides for culture well platelids having extruded features designed to enable the ease of plating orspotting cellular suspensions and other materials either manually or inan automated procedure. Typical cell spotting or plating in anelectrophysiology culture plate or a micro-titer plate can be performedwithout the use of a guide and as shown in FIG. 3. Such an unguidedpipetting technique can be used both manually and via automatedprocesses. As used herein, pipetting refers to the act of deliveringliquid, semiliquid, or semisolid material to a well or surface. The term“pipette” may refer to any tool used to deliver the aforementionedmaterial. As used herein, a pipette tip is the end of the pipette thatdispenses the liquid, semiliquid, or semisolid material to a well orsurface. Use of an unguided pipette 300 can be suitable in applicationswhere the culture plate serves only as a support substrate for cellulargrowth and proliferation, thus precise placement of cells 302 andmaterials is not important. However, in many other cases the cultureplate can be further functionalized with an electroactive area includingwithout limitation biosensors, microelectrodes, microelectronics,impedance electrodes and the like. The biosensors 408 may be fabricatedfrom gold, copper, platinum, nano-textured gold, nano-porous platinum,PEDOT, titanium nitride, indium tin oxide or any other suitableconductive material. For example, using biosensors 408 can enablestimulation and/or recording of cellular activity as well as engage thecell cultures in a variety of applications such as disease in a dishmodels, toxicity screening, phenotypic screening, stem cellcharacterization, and the like. Thus, when functional features areprovided on a culture plate, precise placement or spotting of cells andother materials can become important.

Accordingly and as shown in FIG. 4, it is an aspect of the presentdisclosure to provide for a culture well plate lid 404 that guidespipette 400 for targeted cell plating or spotting. The culture wellplate lid 404 can comprise a plurality of targeting devices orextrusions 406 where each extrusion 406 has openings at its upper andlower ends and where the plurality of extrusions can be configured tomate with a plurality of culture wells present on a culture well plate.The extrusion 406 of the culture plate lid 404 guides the pipette 400 toa precise location (e.g., corresponding to biosensors 408) on theculture well plate thereby facilitating targeted deposition of cellularsuspensions 402 and other materials. In one aspect, these extrusions canapproximate micro to millimeter sized hollow frustocones, hollowpyramidal frustum, tubes, and combinations thereof. In another aspect,the height of the extrusions can range from about 1 mm to about 50 mm.In other embodiments the height of the extrusions may be about 11 mm toabout 12.5 mm.

FIG. 5A shows a lid 404 containing multiple implementations of targetingdevices or extrusions 406. Culture well plate lids can comprise an arrayof extruded micro or milli-scale hollow frustocones or pyramidal orcylindrical frustum without or coupled with a tube at its distal end. Aportion of the extrusion 406, labeled as tube 516 in FIG. 5B, may beshaped to form a collar 518. The collar 518 may engage a ridge on apipette tip, preventing a portion of the pipette from extending beyondthe collar 518. During the dispensing or placement of materials, collar518 maintains a predetermined distance between a pipette tip and abiosensor array. In some embodiments, this predetermined distance may be50 millimeters or less. In other embodiments, the predetermined distancemay be 2.0 millimeters or less. In yet other embodiments, thepredetermined distance may be 0.5 millimeters or less. However, anypredetermined distance may be used, and the location of the collar 518can be designed to provide any suitable distance. Collar 518 may alsomaintain a predetermined distance between the pipette tip and the distalend of the extrusion during the placement of materials. The location ofcollar 518 may influence this predetermined distance. The length ofextrusion may also influence the predetermined distance. Any particularcombination of collar 518 position and extrusion length may be used toprovide the desired distance between the pipette tip and the distal endof the extrusion. As used herein, the distal end of an extrusion is theend located farthest from the rest of the lid 404.

FIG. 5A depicts a variety of types of extrusion profiles to illustratedifferent modalities of the present disclosure and, in practice, theculture well plate lid 404 would likely have a plurality of extrusions406 having a uniform geometry. Although a variety of different extrusionprofiles are shown here, the extrusions 406 are not limited to theseprofiles.

In some embodiments, the culture well plate lid can comprise at leastone extruded micro- or milli-scale hollow frustocone. FIG. 6A is atop-down view of an example targeting device or extrusion 406. FIG. 6Bis a side view of an example extrusion.

In other embodiments, the culture well plate lid can comprise at leastone extruded micro- or milli-scale pyramidal frustum. FIG. 7A is atop-down view of an example extrusion 406, also including collar 518.FIG. 7B is a side view of an example extrusion.

In another embodiment, the culture well plate lid can comprise at leastone extruded micro- or milli-scale tube. FIG. 8A is a top-down view ofan example extrusion 406. FIG. 8B is a side view of an exampleextrusion.

FIG. 9A depicts an example culture well plate lid 404 coupled to amulti-well culture plate 106. The lid 404 includes targeting devices orextrusions 406 that extend into the wells of the culture plate 106. Inthis embodiment, the wells of culture plate 106 are spaced apart fromone another such that there is material filling the space between thewells. In other embodiments, the wells may be closer together withthinner walls separating the wells. FIG. 9A also depicts a biosensor orMEA plate 104 attached to the bottom of culture plate 106. While thisfigure depicts a variety of types of extrusion profiles the lid 404 mayalso have a plurality of extrusions having a uniform geometry.

FIG. 9B shows another example embodiment of a lid 404 fitted into amulti-well culture plate 106. In this example embodiment, the lid 404includes an extrusion 406 that incorporates a collar 518. Collar 518 issized such that a pipette contacts the collar 518 after the pipette hasbeen inserted into the extrusion 406 a sufficient distance. The collar518 maintains a predetermined distance between a pipette tip and abiosensor array. Collar 518 may also maintain a predetermined distancebetween the pipette tip and the distal end of the extrusion during theplacement of materials.

In various example embodiments, extruded frustocones, pyramidal frustum,tubes, and combinations thereof can serve as pipette tip guides forpipetting material and cellular suspensions into an area of interest. Inone embodiment, the area of interest can be a biosensor or a biosensorarray associated with the culture well plate. In one embodiment, thebiosensor array can be an MEA. In other embodiments, pipetted materialscan comprise biomolecular coatings such as, for example and withoutlimitation, laminin, poly-ethylene imine (PEI), poly-ornithine,poly-L-Lysine, fibronectin and the like. In yet other aspects cellularsuspensions can comprise, for example and without limitation, neuralcells, cardiac cells, muscular cells, retinal cells, stem cells, and thelike.

FIGS. 10A-C show additional embodiments of lid 404. As seen in FIG. 10A,a cross section of lid 404, the edge may include a double baffled edge1020 to reduce the amount of fluid lost through evaporation and/ormaintain sterility within the culture plate. FIG. 10B shows a view oflid 404 turned upside down to better depict the features of thisembodiment. In addition to the double baffled edge 1020, the lid mayinclude a friction fit feature 1022. This feature flexes when pressedonto the culture well plate 106 and provides a source of friction thatcan function to keep the lid 404 securely on the well plate 106. It mayalso assist in holding the lid 404 in place while inserting and removingthe pipette tip, further ensuring precise spotting of the cellsuspension onto MEA plate 104. The friction fit feature also serves tolocate the extruded shape to the center of the well. FIG. 10C is a topview of lid 404, further demonstrating the structure of an exampleembodiment of friction fit feature 1022.

FIG. 11 shows a top view of an example embodiment of a culture well 110from a biologic culture plate 106 attached to a monolithic biosensorplate 104. In this example, a droplet of cell suspension 402 ismis-targeted and in contact with both with biosensors 408 and thereference sensors 1124, which may cause misinterpretation of the signalsproduced by the cells. In this embodiment, droplet 402 represents a 5-10microliter drop of material. However, in other embodiments, dropletvolume may be lower than 5 microliters or greater than 10 microliters.

FIG. 12A shows a top view of an example embodiment of a culture well 110with containment devices or features 1228 on the surface of the bottomof the well. Containment devices 1228 are configured to assist in theplacement of materials by physically influencing the positioning of thefluid and concentrating the volume of the cells and other materials tothe biosensors 408. In this embodiment, the containment devices arelevees that are spaced between the biosensors 408 and the referencesensors 1124. In some embodiments, the levees are a minimum distancefrom the nearest biosensor 408, and a minimum distance from the nearestreference sensor 1124. The levees 1228 of the example shown in FIG. 12Aare elongated in shape and follow the curve of the wall of the culturewell 110. However, containment devices 1228 may take any shape and neednot follow the curve of the culture well. For example, the containmentdevices 1228 may be rectangular, spherical, or any other shape that mayassist in the placement of materials. In other aspects, containmentdevices may be moats, weirs, or any other structure designed tophysically influence the positioning of the fluid.

The containment devices or features 1228 may physically influence theplacement of materials by providing an object requiring a force for thematerial to surpass which is less than the surface tension that holdsthe material, such as a cell suspension, together in the center of thewell. The height, shape, and structure of the containment devices 1228may affect the extent to which they influence the positioning of thematerials. For example, taller containment devices 1228 may be able tomore effectively contain the materials, and may also be able to moreeffectively contain a greater quantity of materials, compared to shortercontainment devices 1228. In addition, the number of containment devices1228 can be varied depending on the volume, surface tension, and desiredlocation of the materials being used in the well.

The containment devices 1228 may or may not be electrically active. Insome implementations, the containment devices 1228 may be made from ametal including copper, gold, and so on. However, the containmentdevices 1228 may be made out of any materials that effectivelyaccomplished a goal of, for example, containing the materials within thewell.

FIG. 12B shows a cross section along line A-A′ of FIG. 12A. In thisembodiment, an insulating material 1230 is provided between the variousbiosensors 408, between the biosensors 408 and the containment device1228, and/or between the containment device 1228 and the referencesensor 1124. The insulating material 1230 may be any suitable insulatingmaterial. In one embodiment, the insulating material 1230 is Kapton.

In one embodiment, a portion of the insulating material 1230 is adjacentto a containment device or feature 1228. As shown in FIG. 12B, theinsulating material may slope upward from the bottom of the well plateto abut against one or more containment devices 1228, reference sensors1124, and/or biosensors 408. The slope of the insulating material 1230may physically prevent the spread of the cell suspension 402 toward thereference sensor 1124. In some embodiments, the insulating material 1230further prevents spreading of the cell suspension 402 by hydrophobicinteractions.

In one embodiment, the fabrication of the device includes a first stepof layering an insulating material 1230 over the biosensors 408,reference sensors 1124, and containment devices 1228. The fabricationfurther includes a second step of removing portions of the insulatingmaterial to enable electrical communication between the cell suspension402 and the biosensors. In certain embodiments, the fabrication includesa step of removing portions of the insulating material 1230 from thesurfaces of the containment devices.

In some embodiments, culture well plate lids disclosed herein enablesuperior cell placement in an area of interest. In other embodiments,the culture well plate lids enable lower consumption of expensivereagents and cellular suspensions. In yet other embodiments, thesignal-to-noise ratio during both stimulation and recording in anelectrophysiology culture plate is improved due to placement directly onelectroactive areas of the plate facilitated by the culture well platelid. In some embodiments, the culture well plate lid facilitatesisolation of the cellular suspension from the reference sensors.

The biologic culture plates and lids may be configured to be AmericanNational Standards Institute/Society for Lab Automation and Screening(ANSI/SLAS) compliant. For high-throughput culture systems such aselectrophysiology culture systems, large-area, ANSI/SLAS-complianthigh-throughput culture plate systems plates can be important asindustry standard compliance can provide compatibility with otherhigh-throughput instrumentation such as, for example and withoutlimitation, plate readers, robotics handlers and the like. Suchhigh-throughput culture plates can have well counts of, for example andwithout limitation, 1, 2, 4, 6, 8, 10, 12, 24, 48, 96, 192, 384 or 768,as well as a lid having, for example, 24, 48, 96, 192, 384, 768, or morecorresponding extrusions.

In other aspects, the present disclosure can provide forelectrophysiology culture plates that can be sterilized using simpletreatments to eliminate the risk of cytotoxicity and do not requiresurface preparation (apart from standard biomolecular treatments) forcell culture applications.

In other aspects, the present disclosure provides for a culture wellplate configuration: operable to prevent communication or contaminationbetween adjacent wells. In a further aspect, the lid comprises aplurality of well caps configured to overlie each of the culture wells.In another aspect, each of the plurality of culture wells has the sameheight relative to the peripheral wall of the culture plate.

In other aspects, the present disclosure provides for culture wellplates having culture wells configured to concentrate the volume of thecells/biomolecular treatments deposited specifically on the biosensorarea. In yet other aspects, the present disclosure provides for culturewell plates having culture wells comprising an upper diameter and alower diameter, wherein the upper diameter is greater than the lowerdiameter. In a further aspect, the culture well can circumscribe eithera conical or frustoconical structure on a lower portion of the well.

In another aspect, the biologic culture plate and biosensor platecontain at least one alignment feature configured to define thedirectionality of the plate or align the high-throughput culture wellplate to a die-cut adhesive and the biosensor substrate or both. In afurther aspect, once assembled, the alignment features also align theelectrophysiology culture plate assembly to the docking mechanism andthe high-density connectors located in the electronics unit.

Accordingly, FIGS. 1-12B, and the corresponding text, provide a numberof different devices, systems, methods and mechanisms forhigh-throughput electrophysiology. In addition to the foregoing,implementations described herein can also be described in terms acts andsteps in a method for accomplishing a particular result. For example, amethod comprising at least one of plating, stimulating and recordingdata from a cell culture is described concurrently above with referenceto the components and diagrams of FIGS. 1 through 9.

The present invention can thus be embodied in other specific formswithout departing from its spirit or essential characteristics. Thedescribed aspects are to be considered in all respects only asillustrative and not restrictive. The scope of the invention is,therefore, indicated by the appended claims rather than by the foregoingdescription. All changes that come within the meaning and range ofequivalency of the claims are to be embraced within their scope.

1. A system for targeted placement of materials via a pipette,comprising: a culture plate comprising a plurality of wells; and a lidshaped to removably couple to the culture plate and comprising aplurality of extrusions, wherein at least one of the extrusions extendsinto at least one of the wells, and wherein the at least one of theextrusions is shaped to accept the pipette and direct contents of thepipette into the at least one of the well.
 2. The system of claim 1,wherein the culture plate comprises a biologic culture plate and abiosensor array.
 3. The system of claim 2, wherein the biosensor arraycomprises a biosensor and a reference sensor.
 4. The system of claim 3,wherein the culture plate comprises a containment feature configured toassist in placement of materials on the biosensor.
 5. The system ofclaim 1, wherein the at least one of the extrusions comprises a collarconfigured to prevent a portion of the pipette from extending beyond thecollar.
 6. The system of claim 5, wherein the collar is configured toprevent a pipette tip from protruding more than a predetermined amountbeyond a distal end of the at least one of the extrusions duringplacement of materials via the pipette.
 7. The system of claim 5,wherein the culture plate comprises a biologic culture plate and abiosensor array, and wherein the collar is configured to maintain apredetermined distance between a pipette tip and the biosensor arrayduring placement of materials via the pipette.
 8. The system of claim 1,wherein the at least one of the extrusions is one of a frustoconical,cylindrical, and pyramidal shape.
 9. The system of claim 1, wherein thelid comprises a friction-fit feature, and wherein the friction-fitfeature is configured to secure the lid on the culture plate. 10-21.(canceled)
 22. The system of claim 1, wherein the at least one of theextrusions is configured to guide the pipette to a precise location onthe culture plate.
 23. The system of claim 1, wherein the at least oneof the extrusions is configured to direct the contents of the pipetteinto an area of interest within the at least one of the wells.
 24. Thesystem of claim 2, wherein the contents of the pipette comprise acellular suspension.
 25. The system of claim 2, wherein the biosensorarray is a microelectrode array (MEA).
 26. The system of claim 3,wherein the at least one of the extrusions is configured to direct thecontents of the pipette into an area of interest within the at least oneof the wells, and wherein the area of interest is the biosensor array.27. The system of claim 26, wherein the area of interest within the atleast one of the wells is the biosensor but not the reference sensor.28. The system of claim 4, wherein the containment feature is locatedsuch that it physically influences the positioning of the contents ofthe pipette onto the culture plate.
 29. The system of claim 9, whereinthe friction-fit feature is further configured to flex when in contactwith the culture plate.
 30. The system of claim 9, wherein thefriction-fit feature is further configured to ensure precise spotting ofthe contents of the pipette onto the culture plate.